The primary assimilation of carbon dioxide into organic matter is catalyzed by the enzyme ribulose 1,5-bisphosphate (RuBP) carboxylase/oxygenase (RubisCO). This enzyme also has the capacity to act as an internal monooxygenase and catalyzes the oxygenolytic cleavage of RuBP under suitable conditions. RubisCO is thus a bifunctional protein in which the same polypeptide chain catalyzes the fist step of two competing reactions of cellular metabolism; RubisCO is also the most abundant protein found on earth. This study is directed at two major areas of protein structure and function: the mechanism of complex oligomer assembly and the role of diverse domains in RubisCO catalysis. Recombinant large and small subunits of RubisCO have been purified by separate expression of the structural genes in Escherichia coli and the complex active oligomeric protein has been reconstituted in vitro. In addition, certain residues of the small subunit that affect assembly and influence catalysis have been identified by mutation. Thus, a major goal of the proposed work will focus on the interaction of various subunits and the role certain residues contribute to assembling the oligomer. The role of chaperonin proteins in mediating the proper folding and subsequent assembly of aggregates of large an small subunits will be probed and we will seek to establish the precise pathway of RubisCO oligomer formation. The second major thrust of our research will consider the influence of small subunits on catalysis by large subunits. Again, aided by recombinant DNA procedures, we have developed a convenient system to research this goal. A number of mutant enzymes have been isolated, and will be isolated in the future, to enable us to probe subunit interactions that influence catalysis. The specific domains of the large subunit which contribute to discrimination between the gaseous substrates will also be probed by a combination of molecular manipulation of specific sequences and random selection of strains that express RubisCO with altered properties. Finally, in vivo posttranslational regulation of RubisCO activity will be studied in bacteria and the molecular basis of an apparent reversible modification of RubisCO will be investigated. These studies thus present an excellent opportunity to relate control of cellular metabolism to the function and assembly of an important, yet complex, oligomeric protein.